High incidence of translocations t(11;14)(q13;q32) and t(4;14)(p16;q32) in patients with plasma cell malignancies

Abnormalities involving the 14q32 region are recurrent chromosomal changes in plasma cell malignancies. Recent preliminary molecular analyses found IGH rearrangements in almost 100% of human myeloma cell lines and in 75% of patients. However, no systematic study analyzing the nature of the partner chromosomal regions have been reported thus far. To define the exact incidence of illegitimate IGH rearrangements and the respective incidence of partner genes cloned to date, we analyzed 141 patients with either multiple myeloma (MM, n = 127) or primary plasma cell leukemia (PCL, n = 14) using fluorescence in situ hybridization. The overall incidence of illegitimate recombinations was 57% (80 of 141 patients). Analysis of this incidence according to Durie and Salmon stage, patients' status, i.e., MM versus primary PCL and diagnosis versus relapse, immunoglobulin type and subtype, and beta2-microglobulin value, did not show any correlation. To analyze the nature of the partner chromosomal region, we selected probes specific for the following genes: FGFR3 (4p16), MYC (8q24), CCND1 (11q13), MAF (16q23), and BCL2 (18q21). These probes, combined with differentially labeled 14q32 probes, were used for dual-color fluorescence in situ hybridization on interphase plasma cells. Among the 80 patients with illegitimate IGH rearrangement, we identified 23 IGH-CCND1 fusion cases [i.e., t(11;14)], 17 IGH-FGFR3 fusion cases [i.e., t(4;14)], 3 IGH-MYC fusion cases [i.e., t(8;14)], and only one IGH-MAF fusion case. No IGH-BCL2 fusion case was detected. In 37 of 80 patients, none of these partner genes was involved. Analysis of cases with specific translocations according to their bioclinical features at diagnosis did not show any correlation. This study demonstrated that CCND1 and FGFR3 genes are involved together in about 50% of MM and primary PCL patients with illegitimate IGH rearrangements.     

Lack of ETV6 (TEL) gene rearrangements or p16INK4A/p15INK4B homozygous gene deletions in infant acute lymphoblastic leukemia

Acute lymphoblastic leukemia (ALL) occurring in infants less than 1 year of age differs clinically and biologically from that observed in older children. Cytogenetically, 11q23 translocations are detected in approximately 50% of infant ALLs and fuse the 11q23 gene HRX with a variety of partner chromosomal loci. Overall, HRX rearrangements are detected molecularly in 70-80% of infant ALLs as compared to 5-7% of ALLs arising in older children. Two recently described molecular abnormalities in childhood ALL are ETV6 gene rearrangements and homozygous deletions of p16(INK4A) and/or p15(INK4B). Each of these abnormalities occurs in 15-20% of all childhood ALLs, and neither can be accurately identified by routine cytogenetic analyses. The incidence of these genetic abnormalities and their potential relationship to HRX gene status in infant ALL is unknown. Using Southern blot analyses, we determined ETV6 and p16(INK4A)/p15(INK4B) gene status in a cohort of infant ALLs. No ETV6 rearrangements or homozygous deletions (n=69) or homozygous p16(INK4A) and/or p15(INK4B) gene deletions (n=54) were detected in any of the infant ALLs. Therefore, ETV6 and p16(INK4A)/p15(INK4B) do not play a significant role in the pathogenesis of infant ALL, further emphasizing the distinctive biology of this subset of leukemias.     

A case of primary splenic large cell lymphoma with a t(9;14)(p13;q32)

Appropriately substituted 2,3-dihydro-7H-thiazolo[3,2-a]pyrimidin-7-ones 9-12 and 18 were considered as annulated analogues of HEPT (1-[(2-hydroxyethoxy)methyl]-6-(phenylthio)-thymine), and some of these compounds were also found active against HIV-1, the most active one being 2,3-dihydro-5-[(3,5-dimethylphenyl)methyl]-3-ethoxy-6-ethyl-7H- thiazolo[3,2-a]pyrimidin-7-one (10b). S-Alkylation of 5-alkyl-6-(arylmethyl)-2-thiouracils 1-4 was performed with 2-bromoacetaldehyde acetals to furnish the S-[bis(alkoxy)ethyl] derivatives 5-8 and with allyl bromide to furnish S-allyl derivatives 17. The target compounds 9-12 were obtained by an N1 regioselective intramolecular cyclization reaction of silylated 5-8 using trimethylsilyl trifluoromethanesulfonate (TMS triflate) as the catalyst. Treatment of the S-allyl derivatives 17 with bromine in dry methylene chloride afforded the 3-(bromomethyl) derivatives 18.     

Intragenic mutations of the p16(INK4), p15(INK4B) and p18 genes in primary non-small-cell lung cancers

The p15(INK4B), p16(INK4) and p18 genes are members of the gene family coding for inhibitors of cyclin-dependent kinases 4 and 6. p15(INK4B) and p16(INK4) are located at 9p21, a chromosomal region frequently deleted in many human neoplasms. To examine the role of these 3 genes in lung carcinogenesis, somatic mutations within the genes were analyzed by single-strand conformation polymorphism and DNA sequencing in 71 non-small-cell lung cancer (NSCLC) samples. Six somatic mutations in the p16(INK4) gene and 3 cases with a polymorphic allele were observed. Loss of heterozygosity in the p18 gene was found in 1 sample. We did not find any intragenic mutations in the p15(INK4B) or p18 genes. We conclude that p16(INK4) mutations play a role in the formation of some NSCLCs, whereas the involvement of p15(INK4B) and p18 is uncommon.     

Establishment of a novel myeloma cell line KPMM2 carrying t(3;14)(q21;q32), which proliferates specifically in response to interleukin-6 through an autocrine mechanism

We established a new human myeloma cell line, KPMM2, which proliferates specifically in response to IL-6 via an autocrine mechanism. The proliferative response of KPMM2 cells to exogenous IL-6 was significantly stimulated in a dose-dependent manner. The growth was markedly inhibited by an anti-IL-6 mAb and an anti-IL-6 receptor (IL-6R) mAb in a dose-dependent manner. KPMM2 cells expressed IL-6 and IL-6R mRNA by RT-PCR. Flow cytometric analysis showed cell surface expression of IL-6R. IL-6 protein was detected in the culture supernatant by ELISA. IL-11, oncostatin M and leukemia inhibitory factor had no effect on the proliferation of KPMM2 cells although interferon-alpha and interferon-gamma inhibited the growth. Furthermore, KPMM2 cells bore a t(3;14)(q21;q32) translocation and this finding is of potential interest for future studies in the light of the nuclear protein BM28 (CDCL1, for cdc-like 1) mapped on 3q21, which plays an important role in the cell cycle. In this report, we demonstrated completely an IL-6-dependent autocrine growth mechanism in KPMM2 cell line. This cell line may be useful to investigate the pathogenesis of multiple myeloma and to evaluate the therapeutic potential of IL-6 blocking agents in vitro and in vivo.     

A der(22)t(9;22)(p13;q11) in bone marrow cells of a patient with multiple myeloma: a variant of the t(9;14)(p13;q32) associated with a subset of lymphoplasmacytoid lymphoma

An adult cattle egret (Bubulcus ibis) caught in Kobe of Hyogo Prefecture, Japan, in December 1995 died of a severe infection associated with the trematode parasite Pegosomum sp. At necropsy, 22 trematode parasites were found in the lumen of the bile duct, and the duct wall was markedly thickened. Histopathologically, severe cholangitis and cholecystitis were observed in close association with the parasites in the bile duct. Severe Pegosomum sp. infection may be one of the factors contributing to the mortality of wild cattle egrets. This is the first reported case of the genus Pegosomum infection in wild birds of Japan.     

Chromosome 18 breakpoint in t(11;18)(q21;q21) translocation associated with MALT lymphoma is proximal to BCL2 and distal to DCC

OBJECTIVE: To identify independent predictors of intracranial hemorrhage (ICH) during neonatal extracorporeal membrane oxygenation (ECMO). STUDY DESIGN: This retrospective cohort consisted of all neonates who did not have an ICH before treatment with ECMO identified in the Extracorporeal Life Support Organization Registry from 1992 to 1995 (n = 4550). Multiple logistic regression analysis was used to identify factors independently correlated with ICH and to develop a model that could be used to predict the risk of ICH in neonates treated with ECMO. RESULTS: ICH was identified in 9.9% of patients. The factors associated with ICH remaining after adjusting for other significant variables (P <.01) were gestational age (GA) <34 weeks (odds ratio [OR] 12.1, 95% confidence intervals [CI] [6.6, 22]), GA 34 to <36 weeks (OR 4.1, CI [2.9, 5.8]), GA 36 to <38 weeks (OR 2.1, CI [1.6, 2.8]) primary diagnosis of sepsis (OR 1.8, CI [1.4, 2.3]), epinephrine use (OR 1.9, CI [1.5, 2.5]), coagulopathy (OR 1. 6, CI [1.1, 2.2]), arterial pH <7.0 (OR 2.5, CI [1.6, 3.9]), and arterial pH 7.0 to <7.2 (OR 1.8 CI [1.3, 2.5]). ICH rates for neonates receiving venovenous versus venoarterial ECMO and for those treated with or without cephalic jugular venous drainage were not significantly different. CONCLUSIONS: Gestational age, acidosis, sepsis, coagulopathy, and treatment with epinephrine are major independent factors associated with ICH in neonates treated with ECMO. In particular, GA <34 weeks remains a major barrier for use of current ECMO technologies.     

Homozygous deletions of methylthioadenosine phosphorylase (MTAP) are more frequent than p16INK4A (CDKN2) homozygous deletions in primary non-small cell lung cancers (NSCLC)

Homozygous deletions of the tumor suppressor gene p16INK4A and deficiency of methylthioadenosine phosphorylase (MTAP), both located on chromosome 9p21, have been independently reported in non-small cell lung cancer (NSCLC). To determine the frequency of co-deletion of these two genes, we investigated 50 samples of primary NSCLC using a quantitative PCR-ELISA. All specimens were fixed in formalin, paraffin embedded and stored until assayed. Histologic subtypes included 25 adenocarcinomas (50%), 21 squamous cell carcinomas (42%) and four large cell carcinomas (8%). Homozygous deletions of MTAP exon 8 could be detected in 19 of 50 NSCLC samples (38%). Adenocarcinoma (11 of 25, 44%) showed a higher deletion frequency than squamous cell carcinoma (six of 21, 29%). In contrast, homozygous p16INK4A deletions were detected in only nine of 50 (18%) samples using specific primers for p16INK4A exon 1alpha. No difference between the histological subtypes and p16INK4A deletion frequency was observed. We further investigated the ten samples with MTAP deletions but intact p16INK4A exon 1alpha with primers specific for p16INK4A exon 3, the exon nearest to MTAP exon 8. Interestingly, none of the ten samples had deletion of the p16INK4A exon 3 coding region. Fine mapping analysis performed in ten samples showed a frequent breakpoint between MTAP exon 4 and exon 5. In addition, p16 protein expression could not be detected in five out of six samples with intact p16 but deleted MTAP locus. These data show a high frequency of homozygous MTAP deletions in NSCLC which is associated with detectable co-deletion of p16INK4A in only half of the cases. This result suggests the existence either of another tumor suppressor gene telomeric of p16INK4A or of deletions involving 3'-untranslated (3'-UTR) regulatory regions of p16INK4A that can interfere with its expression or function.     

Lack of germ-line mutations of CDK4, p16(INK4A), and p15(INK4B) in families with glioma

Gliomas are tumors of the central nervous system that may be inherited in some patients. The gene(s) responsible for the clustering of gliomas in families have not yet been identified. Molecular studies of sporadic high-grade gliomas have revealed mutations or deletions of the genes encoding the protein kinase inhibitors p16(INK4A) and p15(INK4B) in a large proportion of tumors. Moreover, those tumors without deletions frequently display gene amplification and/or over-expression of mRNA encoding the protein kinase cdk4. We hypothesized that germ-line mutations in the p16(INK4A), p15(INK4B), or CDK4 genes might contribute to some cases of familial gliomas. To address this issue, we analyzed 36 kindreds with a predisposition to glial tumors. Genomic DNA from index members of these families was screened by PCR-single-strand conformational polymorphism analysis. We did not detect any functional mutations in the p16(INK4A), p15(INK4B), or CDK4 genes, although two individuals did have a previously described A140T polymorphism in p16(INK4A). Thus, despite the association between the sporadic forms of high-grade glioma and abnormalities of p16(INK4A), p15(INK4B), or CDK4, we found no evidence that germ-line mutations in the coding region of these three genes predispose to inherited glial tumors.     

Prenatal diagnosis of a der(X)t(X;15)(p22.2;q11.2) inherited from a maternal translocation X;15

During a 15-y period, 48 children were treated for Wilms' tumour (WT). Seven of them were < 1 y of age at diagnosis. One child presented with non-traumatic haematuria, but in all the other children WT was revealed as a palpable abdominal mass at routine examination or investigation due to another disease. The four children under 6 months of age at diagnosis were primarily operated upon; the others received preoperative chemotherapy. Two children had chromosomal aberrations in the WT tumour specimen. The follow-up revealed that postoperatively six children are healthy with no evidence of relapse from WT, but one child had a contralateral relapse successfully enucleated. The clinical behaviour and management of WT in infants differ compared with that in older children. The diagnosis may be uncertain and it can be difficult to distinguish between malignant and non-malignant lesions. It is essential to realize the possibility of WT, even in children < 1 y of age.     

Translocation (11;14)(q13;q32) and overexpression of cyclin D1 protein in a CD23-positive low-grade B-cell neoplasm

BACKGROUND: Platinum compounds are the most active drugs in ovarian cancer treatment; cisplatin and carboplatin demonstrated similar efficacies but different toxicity profiles. Paclitaxel combined with cisplatin as first-line treatment improved overall survival when compared to a cisplatin-cyclophosphamide combination, but generated higher rates of neutropenia, febrile neutropenia and neurotoxicity. The paclitaxel-carboplatin combination may be better tolerated than cisplatin-paclitaxel. DESIGN: The objective of the present study was to assess the efficacy and safety of the combination of paclitaxel and carboplatin in previously treated advanced ovarian cancer patients. PATIENTS AND METHODS: During or after platinum-based chemotherapy, 73 patients with progressive advanced epithelial ovarian carcinoma were enrolled to receive every four weeks a three-hour infusion of paclitaxel 175 mg/m2 followed by a 30-minute carboplatin infusion. The carboplatin dose was calculated to obtain the recommended area concentration-versus-time under the curve of 5 mg x ml-1 x min. RESULTS: Toxicity and response could be evaluated for 72 and 62 patients, respectively. Eleven complete and 15 partial responses gave an overall response rate of 42% (95% CI: 30%-54%). Response rates for platinum-refractory patients and those with early (> or = 3 and < 12 months) and late (> 12 months) relapses were 24%, 33% and 70%, respectively. The respective median response duration, the median progression-free survival and median overall survival were 8, 6 and 14 months. Myelosuppression was the most frequent and severe toxicity. Grade 3 and 4 neutropenia occurred, respectively in 30% and 23% of the cycles; 6% of the cycles benefited from medullary growth factors. Only one episode of febrile neutropenia was observed. Grade 3 and 4 thrombocytopenia occurred, respectively during 3% and 1% of the cycles. Alopecia was frequent. Transient peripheral neuropathy developed in 47% of patients but was severe in only one patient. One early death was attributed to progressive disease and possibly to therapy. CONCLUSION: This combined paclitaxel-carboplatin therapy is effective and can be safely administered to ovarian cancer patients who relapse after one or two regimens of platinum-based chemotherapy.     

The LMO1 and LDB1 proteins interact in human T cell acute leukaemia with the chromosomal translocation t(11;14)(p15;q11)

The ectopic expression of LMO1 or LMO2 in T cell acute leukaemias resulting from chromosomal translocations t(11;14)(p15;qll) or t(11;14)(p13;q11) respectively in a causal factor in tumorigenesis. LMO1 has been found as a heterodimer with a 46 Kd protein in a T cell line derived from a childhood T-acute leukaemia. This 46 Kd protein is the LIM-binding protein LDB1/NLI. The latter is a phosphoprotein and binds to LMO1 in its phosphorylated state and essentially all the LMO1 and LDB1 protein in the T cell line is part of the complex. Therefore, the LMO1-LDB1 interaction is likely to be involved in tumorigenesis after LMO1 is ectopically expressed following chromosomal translocation in T cells prior to development of acute leukaemias.     

Establishment of a novel human myeloid leukaemia cell line (FKH-1) with t(6;9)(p23;q34) and the expression of dek-can chimaeric transcript

Calcein-labeled B16 melanoma (B16M) cells were injected intraportally, and in vivo video microscopy was used to study the distribution and damage of cancer cells arrested in the liver microvasculature over a period of 4 hours. The contribution of glutathione (GSH)-dependent antioxidant machinery to the possible oxidative stress-resistance mechanism of B16M cell was determined by in vitro incubation with the selective inhibitor of GSH synthesis L-buthionine (S,R)-sulphoximine (BSO) before B16M cell injection in untreated and 0.5-mg/kg lipopolysaccharide (LPS)-treated mice. In addition, untreated and LPS-treated isolated syngeneic hepatic sinusoidal endothelial cells (HSE) were used to determine in vitro their specific contribution to B16M cell damage. Trauma inherent to intrasinusoidal lodgement damaged 35% of B16M cells in both normal and LPS-treated mouse liver. The rest of the arrested B16M cells remained intact in normal liver for at least 4 hours, although their damaged cell percentage significantly (P < .05) increased since the second hour in normal mice injected with BSO-treated cells and since the first hour in LPS-treated mice given untreated cells. Recombinant human interleukin-1 receptor antagonist (rHuIL-1-Ra) given to mice 15 minutes before LPS significantly (P < .05) abrogated B16M cell damage. On the other hand, 40% of the B16M cells co-cultured with unstimulated HSE and 70% of the co-cultured with LPS-treated HSE became sensitive to endothelial cell-mediated damage after BSO treatment. These results demonstrate that a high intracellular level of GSH protects B16M cells from possible in vivo and in vitro sinusoidal cell-mediated oxidative stress, contributing to the mechanism of metastatic cell survival within the hepatic microvasculature.